- Phosphate limitation caused significant changes in the fatty acid and lipid composition of Monodus subterraneus. With decreasing phosphate availability from 175 to 52.5, 17.5 and 0 μM (K2HPO4), the proportion of the major VLC-PUFA, eicosapentaenoic acid (EPA), gradually decreased from 28.2 to 20.8, 19.4 and 15.5 mol% (of total fatty acids), respectively. The cellular total lipid content of starved cells increased, mainly due to the dramatic increase in triacylglycerols (TAG) levels. Among polar lipids, cellular contents of digalactosyldiacylglycerol (DGDG) and diacylglyceroltrimethylhomoserine (DGTS) increased sharply from 0.29 and 0.19 to 0.60 and 0.38 fg cell−1, respectively, while that of monogalactosyldiacylglycerol (MGDG) was not significantly changed. In the absence of phosphate, the proportion of phospholipids was significantly reduced from 8.3% to 1.4% of total lipids, and the proportion of triacylglycerols (TAG) increased from 6.5% up to 39.3% of total lipids. The share of MGDG was substantially reduced, from 35.7% to 13.3%, while that of DGDG and DGTS reduced less from 18.3% to 15.1%, and 12.2% to 8.6%, respectively. The most distinctive change in the fatty acid composition was noted in that of DGDG, where the proportion of EPA, located exclusively at the sn-1 position, increased from 11.3% to 21.5% at the expense of 16:0, 16:1 and 18:1. In MGDG, however, the proportion of EPA did not change appreciably. In contrast to higher plants, DGDG accumulated under P-deprivation in M. subterraneus, did not resemble PC and the positional distribution of its fatty acids was not altered, preserving the C20/C16 structure of its molecular species. We suggest that under phosphate starvation DGTS is a likely source of C20 acyl groups that can be exported to the sn-1 position of DGDG and can partially compensate for the decrease in PE, the apparent source of C20 acyl-containing diacylglycerols in this alga. Moreover, accumulation of non-esterified 18:0 indicates that no polar lipid can replace PC, which appears to be the only lipid capable of C18 desaturation in this alga.